Targets for Receptor-Mediated Control of Therapeutic Biodistribution and Efficacy

Abstract

Disclosed herein include novel blood-brain barrier (BBB)-crossing receptors on the BBB interface, targeting peptides and derivatives thereof capable of binding to the novel receptors, and related methods of using the receptors to increase the permeability of the BBB and to deliver an agent to a nervous system (e.g., CNS). In some embodiments, the BBB-crossing receptor is LRP6. Disclosed herein also include recombinant adeno-associated viruses (rAAVs) with increased specificity and transduction efficiency across the BBB and related compositions and methods of treating various diseases and conditions.

1 . A method of increasing permeability of the blood brain barrier, comprising:
 providing a targeting peptide capable of binding to low density lipoprotein receptor related protein 6 (LRP6), thereby increasing permeability of the blood brain barrier, wherein the targeting peptide binds YWTD domain 1 and/or domain 2 of LRP6.   
     
     
         2 . (canceled) 
     
     
         3 . The method of  claim 1 , wherein the permeability of the blood brain barrier is increased by at least 25%, 50%, 75%, 100%, or more as compared to the absence of the targeting peptide. 
     
     
         4 . A method of delivering a payload to a nervous system of a subject, the method comprising:
 providing a targeting peptide capable of binding to low density lipoprotein receptor related protein 6 (LRP6) or a derivative thereof, wherein the targeting peptide is part of a delivery system, and wherein the delivery system comprises the payload to be delivered to the nervous system; and   administering the delivery system to the subject.   
     
     
         5 . The method of  claim 4 , wherein the delivery system comprises nanoparticles, nanotubes, nanowires, dendrimers, liposomes, ethosomes and aquasomes, polymersomes and niosomes, foams, hydrogels, cubosomes, quantum dots, exosomes, macrophages, and any combination thereof; or
 wherein the delivery system comprises a viral vector or a non-viral vector, wherein the targeting peptide enhances the binding affinity of the viral vector or the non-viral vector to LRP6.   
     
     
         6 . (canceled) 
     
     
         7 . (canceled) 
     
     
         8 . The method of  claim 5 , wherein the viral vector comprises an AAV vector; wherein the targeting peptide is part of a capsid protein of an AAV vector. 
     
     
         9 . The method of  claim 8 , wherein the AAV vector is a vector selected from the group consisting of AAV1, AAV2, AAV3, AAV3b, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV-DJ, human isolate hu.31, human isolate hu.32, rhesus isolate rh.8, rhesus isolate rh.10, and a variant thereof. 
     
     
         10 . The method of  claim 5 , wherein the non-viral vector comprises lipid-based nanoparticles, polymeric nanoparticles, inorganic nanoparticles, surfactant-based emulsions, nanowires, silica nanoparticles, peptide or protein-based particles, lipid-polymer particles, nanolipoprotein particles, and combinations thereof. 
     
     
         11 . The method of  claim 4 , wherein the payload to be delivered to a nervous system is a biological molecule, a non-biological molecule, or a combination thereof; wherein the biological molecule is selected from the group consisting of a nucleic acid sequence, a protein, a peptide, a lipid, a polysaccharide, and any combination thereof. 
     
     
         12 . The method of  claim 4 , wherein the payload is a therapeutic molecule. 
     
     
         13 . The method of  claim 11 , wherein the nucleic acid sequence to be delivered to a nervous system comprises one or more of:
 a) a sequence encoding a trophic factor, a growth factor, or other soluble factors that might be released from the transduced cells and affect the survival or function of that cell and/or surrounding cells;   b) a DNA that restores protein function to humans or animals harboring a genetic mutation(s) in that gene;   c) a DNA that encodes a protein that can be used to control or alter the activity or state of a cell;   d) a DNA that encodes a protein or a nucleic acid used for assessing the state of a cell;   e) a DNA and/or associated guide RNA for performing genomic engineering;   f) a sequence for genome editing via homologous recombination;   g) a DNA sequence encoding a therapeutic RNA;   h) an shRNA or an artificial miRNA delivery system; or   i) a DNA sequence that influences the splicing of an endogenous gene.   
     
     
         14 . The method of  claim 4 , wherein the LRP6 is a mouse LRP6; wherein the LRP6 has an amino acid sequence having at least 80% sequence identity to an amino acid sequence of SEQ ID NO: 29; or wherein the LRP6 is a macaque LRP6; wherein the LRP6 has an amino acid sequence having at least 80% sequence identity to an amino acid sequence of SEQ ID NO: 30; or wherein the LRP6 is a human LRP6; wherein the LRP6 has an amino acid sequence having at least 80% sequence identity to an amino acid sequence of SEQ ID NO: 31. 
     
     
         15 . (canceled) 
     
     
         16 . (canceled) 
     
     
         17 . The method of  claim 4 , wherein upon binding the targeting peptide is capable of interacting with (1) one or more positions functionally equivalent to R28, G158, E159, W183, A201, K202, or H226 in LRP6 having an amino acid sequence of SEQ ID NO: 31; or (2) one or more positions functionally equivalent to S96, S114, E115, R141, W157, W183, or W242 in LRP6 having an amino acid sequence of SEQ ID NO: 31. 
     
     
         18 . The method of  claim 9 , wherein the targeting peptide is inserted between two adjacent amino acids in AA587-594 of SEQ ID NO: 11 of the AAV9 vector or functional equivalents of AA587-594 in an amino acid sequence at least 80% identical to SEQ ID NO: 11; or wherein the targeting peptide is inserted between AA588-589 of SEQ ID NO: 11 of the AAV9 vector or functional equivalents of AA588-589 in an amino acid sequence at least 80% identical to SEQ ID NO: 11. 
     
     
         19 . (canceled) 
     
     
         20 . The method of  claim 8 , wherein the AAV vector is conjugated to a nanoparticle, a second molecule, or a combination thereof. 
     
     
         21 . The method of  claim 4 , wherein the administration is a systemic administration. 
     
     
         22 . The method of  claim 4 , wherein the subject is a human and wherein the subject is a subject suffering from or at a risk to develop one or more of chronic pain, Friedreich's ataxia, Huntington's disease (HD), Alzheimer's disease (AD), Parkinson's disease (PD), Amyotrophic lateral sclerosis (ALS), spinal muscular atrophy types I and II (SMA I and II), Friedreich's Ataxia (FA), Spinocerebellar ataxia, multiple sclerosis (MS), chronic traumatic encephalopathy (CTE), HIV-1 associated dementia, or lysosomal storage disorders that involve cells within the CNS; wherein the lysosomal storage disorder that involve cells within the CNS is Krabbe disease, Sandhoff disease, Tay-Sachs, Gaucher disease (Type I, II or III), Niemann-Pick disease (NPC1 or NPC2 deficiency), Hurler syndrome, Pompe Disease, or Batten disease. 
     
     
         23 . (canceled) 
     
     
         24 . The method of  claim 4 , wherein the subject is a subject suffering from, at risk to develop, or has suffered from a stroke, traumatic brain injury, epilepsy, or spinal cord injury. 
     
     
         25 .- 49 . (canceled) 
     
     
         50 . A method of designing a targeting peptide having specificity to low density lipoprotein receptor related protein 6 (LRP6), comprising:
 generating in silico one or more targeting peptides each capable of interacting with (1) one or more positions functionally equivalent to R28, G158, E159, W183, A201, K202, or H226 in LRP6 having an amino acid sequence of SEQ ID NO: 31; or (2) one or more positions functionally equivalent to S96, S114, E115, R141, W157, W183, or W242 in LRP6 having an amino acid sequence of SEQ ID NO: 31.   
     
     
         51 . The method of  claim 50 , wherein generating in silico the one or more targeting peptides comprises:
 generating in silico a plurality of candidate peptides;   performing computer-assisted docking simulations for each of the plurality of candidate peptides binding to LRP6; and   analyzing the structure of LRP6 binding to one or more of the plurality of candidate peptides to identify one or more targeting peptides capable of interacting with (1) one or more positions functionally equivalent to R28, G158, E159, W183, A201, K202, or H226 in LRP6 having an amino acid sequence of SEQ ID NO: 31; or (2) one or more positions functionally equivalent to S96, S114, E115, R141, W157, W183, or W242 in LRP6 having an amino acid sequence of SEQ ID NO: 31.   
     
     
         52 . The method of  claim 50  comprising:
 obtaining a binding score for each of the plurality of candidate peptides binding to LRP6; and 
 selecting one or more of the plurality of candidate peptides having a binding score above a threshold value as a targeting peptide having specificity to LRP6; wherein the method